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首页> 外文期刊>Journal of cell biology >Sorting of mannose 6-phosphate receptors and lysosomal membrane proteins in endocytic vesicles.
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Sorting of mannose 6-phosphate receptors and lysosomal membrane proteins in endocytic vesicles.

机译:内吞囊泡中的甘露糖6-磷酸受体和溶酶体膜蛋白的分类。

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摘要

The intracellular distributions of the cation-independent mannose 6-phosphate receptor (MPR) and a 120-kD lysosomal membrane glycoprotein (lgp120) were studied in rat hepatoma cells. Using quantitative immunogold cytochemistry we found 10% of the cell's MPR located at the cell surface. In contrast, lgp120 was not detectable at the plasma membrane. Intracellularly, MPR mainly occurred in the trans-Golgi reticulum (TGR) and endosomes. lgp120, on the other hand, was confined to endosomes and lysosomes. MPR was present in both endosomal tubules and vacuoles, whereas lgp120 was confined to the endosomal vacuoles. In cells incubated for 5-60 min with the endocytic tracer cationized ferritin, four categories of endocytic vacuoles could be discerned, i.e., vacuoles designated MPR+/lgp120-, MPR+/lgp120+, MPR-/lgp120+, and vacuoles nonimmunolabeled for MPR and lgp120. Tracer first reached MPR+/lgp120-, then MPR+/lgp120+, and finally MPR-/lgp120+ vacuoles, which are assumed to represent lysosomes. To study the kinetics of appearance of endocytic tracers in MPR-and/or lgp120-containing pools in greater detail, cells were allowed to endocytose horse-radish peroxidase (HRP) for 5-90 min. The reduction in detectability of MPR and lgp120 antigenicity on Western blots, due to treatment of cell homogenates with 3'3-diaminobenzidine, was followed in time. We found that HRP reached the entire accessible pool of MPR almost immediately after internalization of the tracer, while prolonged periods of time were required for HRP to maximally access lgp120. The combined data suggest that MPR+/lgp120+ vacuoles are endocytic vacuoles, intermediate between MPR+/lgp120-endosomes and MPR-/lgp120+ lysosomes, and represent the site where MPR is sorted from lgp120 destined for lysosomes. We propose that MPR is sorted from lgp120 by selective lateral distribution of the receptor into the tubules of this compartment, resulting in the retention of lgp120 in the vacuoles and the net transport of lgp120 to lysosomes.
机译:在大鼠肝癌细胞中研究了不依赖阳离子的甘露糖6磷酸受体(MPR)和120 kD溶酶体膜糖蛋白(lgp120)的细胞内分布。使用定量免疫金细胞化学,我们发现10%的细胞MPR位于细胞表面。相反,在质膜上不能检测到lgp120。在细胞内,MPR主要发生在反式高尔基网(TGR)和内体中。另一方面,lgp120仅限于内体和溶酶体。 MPR同时存在于内体小管和液泡中,而lgp120则局限于内体液泡中。在与内吞示踪剂阳离子化铁蛋白孵育5-60分钟的细胞中,可以识别出四类内吞空泡,即称为MPR + / lgp120-,MPR + / lgp120 +,MP​​R- / lgp120 +的液泡以及未免疫标记MPR和lgp120的液泡。示踪剂首先到达MPR + / lgp120-,然后到达MPR + / lgp120 +,最后到达MPR- / lgp120 +液泡,假定它们代表了溶酶体。为了更详细地研究内含示踪剂在含有MPR和/或lgp120的池中的出现动力学,允许细胞内吞辣根过氧化物酶(HRP)5-90分钟。及时追踪由于3'3-二氨基联苯胺处理细胞匀浆而导致的蛋白质印迹中MPR和lgp120抗原性的可检测性下降。我们发现,在跟踪程序内部化之后,HRP几乎立即到达了MPR的整个可访问池,而HRP最长访问lgp120则需要更长的时间。合并的数据表明,MPR + / lgp120 +液泡是内吞性液泡,位于MPR + / lgp120-内体和MPR- / lgp120 +溶酶体之间,代表从预定用于溶酶体的lgp120分离MPR的位点。我们建议通过受体的选择性侧向分布进入该小室的小管,从lgp120筛选出MPR,从而将lgp120保留在液泡中,并将lgp120净转运至溶酶体。

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